In recent years piperazine derivatives have emerged as a new class of designer drugs, gaining popularity especially among young people in the dance music scene, where they are commonly known as ‘party pills’. They are often marketed as ‘natural’ or ‘herbal’ products, but are in fact entirely synthetic chemicals. Coupled with their ease of availability and varying legal status across the globe, this can create the misconception that these drugs are safe and without the risks commonly associated with traditional street drugs. Piperazine derivatives are usually found in illicit dosage forms as either tablets or capsules, but loose powders and, more rarely, solutions also occur. The tablets often carry logos similar to those seen on ecstasy tablets and they are often misleadingly sold as ecstasy or as apparent ‘safer’ and ‘legal’ alternatives.
Their name derives from the piperazine heterocycle, which is a common feature, and they can be divided into two sub-classes; the benzylpiperazines such as 1-benzylpiperazine (BZP) itself, and the phenylpiperazines, which include 1-(3-chlorophenyl)piperazine (mCPP), 1-(4-methoxyphenyl)piperazine (MeOPP), and 1-(3-Trifluoromethylphenyl)piperazine (TFMPP). Synonyms for BZP include 1-(phenylmethyl)piperazine, 1-Benzyl-1,4-diazacyclohexane, and N-benzylpiperazine. Street names include A2, Bliss, Charge, Frenzy, Herbal ecstasy, Legal E and Legal X.
1-Benzylpiperazine is metabolized in mammals by ring hydroxylation of the aromatic ring, N-dealkylation, O-methylation, and conjugation. Tsutsumi et al. (2006) suggested that p-hydroxy-BZP (p-OH-BZP) is the most relevant target analyte for use in forensic toxicological and clinical analysis of BZP intake, since it retains the structural identity of the parent molecule. Their study, conducted on Wistar rats, found that urinary levels of the parent BZP peaked at around 4 hrs post-dosing, while the levels of the OH-BZP metabolites, p-OH-BZP and m-hydroxy-BZP (m-OH-BZP)-(4-OH-BZP and 3-OH-BZP, respectively), reached maximum levels around 8hrs post-dosing. After the peak, they found that levels of BZP and its OH metabolites in urine dropped rapidly, with BZP becoming undetectable at 48hrs post dosing, while the p-OH-BZP and m-OH-BZP were detectable up to 48hrs post-dosing. Of the doses given in the study, 6.7% was excreted unchanged as the parent BZP molecule, 25% as p-OH-BZP (the most abundant primary metabolite), and 2% as m-OH-BZP. BZP can be detected in the plasma within 30 min of an oral dose of BZP hydrochloride, with peak plasma concentrations occurring between 60 min and 90 min post ingestion (Anita et al., 2009). BZP has an elimination half life of 5-6 hrs and elimination is complete by around 44 hrs.
Analytical methods which have been used to detect or determine piperazine derivatives include gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS) (Staack & Maurer, 2003; Antia et al., 2010). Peters et al. (2010) present a review of analytical toxicology of emerging drugs of abuse, including piperazines. A disadvantage of mass spectrometry based methods of detection is that they require expensive equipment and highly-trained staff.
Immunoassays are known in the art as relatively cost effective, simplistic, and rapid alternatives to mass-spectrometry-based analysis. A commercially available 3,4-methylenedioxy-N-methylamphetamine (MDMA) immunoassay, the EMIT® II Plus Ecstasy Assay (SYVA/Dade Behring), has been shown to have cross-reactivity to BZP, but concentrations of 3,000,000 ng/mL were needed for a positive result (Logan et al., 2010). Neogen Corporation (Lansing Mich., USA) have a specific benzylpiperazine ELISA kit available (Neogen Forensic Reference Brochure). However, no data are provided for the detection of metabolites such as the important 4-OH metabolite (p-OH-BZP) nor is it indicated that the kit has the ability to detect these metabolites. Therefore, there remains a need for a benzylpiperazine assay, which is not only sensitive to the parent molecule but that can also detect key metabolites to enable improvements in the forensic toxicological and clinical analysis of the intake of this designer drug.